Analytical method and apparatus

ABSTRACT

The presence of volatile material, such as ethanol, in biological fluids, such as blood, is determined qualitatively and semiqualitatively by placing a drop of the biological fluid on a porous support so as to vastly extend the surface area of the sample of biological fluid. The expanded sample is confined in a small vapor-tight chamber with a liquid colorimetric agent. Changes in the colorimetric agent are noted.

United States Patent [1 1 Graham June 10, 1975 ANALYTICAL METHOD ANDAPPARATUS [76] Inventor: J. Wallace Graham, 1501 N. Bundy Dr., LosAngeles, Calif. 90049 [22] Filed: Aug. 3, 1972 21 Appl. No.: 277,775

[52] US. Cl. 23/230 B; 206/205 [51] Int. Cl ..G01n 31/22; G0ln 33/16[58] Field of Search 23/230 B, 259, 292, 232 R,

23/258.5, 230 R, 253 TP, 253 R; 206/1 R [56] References Cited UNITEDSTATES PATENTS 3,150,932 9/1964 Ogg 23/259 X 3,389,967 6/1968l-lrabinski 23/230 R Primary Examiner-Robert M. Reese [57] ABSTRACT Thepresence of volatile material, such as ethanol, in biological fluids,such as blood, is determined qualitatively and semiqualitatively byplacing a drop of the biological fluid on a porous support so as tovastly extend the surface area of the sample of biological fluid. Theexpanded sample is confined in a small vaportight chamber with a liquidcolorimetric agent. Changes in the colorimetric agent are noted.

13 Claims, 6 Drawing Figures ANALYTICAL METHOD AND APPARATUS Often, whena patient is admitted to an emergency ward, that patient is unconscious.It often appears, probable that some chemical agent is responsible forthe patients condition; however. it is usually difficult, if notimpossible, to determine which chemical agent or combination of chemicalagents is responsible for the patients condition. The treatmentnecessary to save the patients life is dependent upon determiningquickly and accurately which chemical agents are involved andapproximately how much of the chemical agent the patient has consumed.Speed, simplicity, and accuracy are essential in making the qualitativeand semiquantitative determination of the chemical agent that isresponsible for the patients unconscious condition.

Many of the chemical agents that frequently contribute to theunconsciousness of persons who are admitted to emergency wards arevolatile. Probably the most common of these volatile chemicals isethanol.

Previous solutions to the problem of qualitative and semiquantitativeanalysis of chemical agents is biological fluids have generally requiredtoo much time or have been too complicated for rapid use under emergencyconditions with an unconscious patient.

According to the present invention, these and other difficulties of theprior attempts to solve the problem of quickly analyzing biologicalfluids under emergency conditions for volatile chemical agents have beenovercome, and the advantages of a rapid, simple, and accuratequalitative and semiquantitative analysis have been achieved.

The present invention provides a quick, simple, and accurate procedureand the physical kit for carrying out that procedure for determining thepresence of volatile materials; such as ethanol, methanol, andisopropanol, in biological fluids, such as blood and urine. A quickqualitative and semiquantitative indication of the volatiles in thebiological fluid is obtained within minutes of the time that the sampleis withdrawn from the patient.

In general, a drop or two of biological fluid that is suspected ofcontaining a volatile material is placed on a porous support member thatvastly extends the surface of the biological fluids sample so as topromote the rapid volatilization of the volatile chemical agent. Theexpanded sample is placed in a small vapor-tight chamber so that thevolatilized chemical agent is confined to a small vapor space. Ananalytic reagent is also placed in the same chamber so that thevolatilized chemical agent in the vapor phase contacts the analyticreagent. The analytic reagent is chosen so that it gives a detectableindication responsive both to the presence and concentration of thevolatile chemical agent in the vapor phase. Preferably, the analyticreagent is liquid, and the vapor-tight reaction chamber is proportionedso that the liquid analytic reagent presents a large surface area to thevapor phase space that is shared in common with the expanded sample ofbiological fluid. Preferably, the analytic reagent is a colorimetricreagent that changes color responsive to the presence of the volatilematerial being tested for. Preferably, the color change takes place at arate proportionate to the concentration of the volatile material in thesample of biological fluid.

In the drawings there is illustrated:

FIG. 1, a broken side elevation view of an analytic package according tothe present invention;

FIG. 2, a cross-sectional plan view taken along line 2-2 in FIG. 1;

FIG. 3, a plan view similar to FIG. 2 except that the porous member isshown in the expanded configuratron;

FIG. 4, a side elevation of a further embodiment of the analytic packageshown in FIG. 1, including an optical filter;

FIG. 5, a cross-sectional side elevation of the embodiment shown in FIG.4 taken along 55 in FIG. 4; and

FIG. 6, a partially broken side elevation view ofa further embodiment ofan analytic package according to the present invention.

Referring particularly to the drawings) there is illustrated ananalytical package indicated generally at 10, comprising a reactionchamber 12 and a sample structure 22. Reaction chamber 12 is defined bya generally cylindrical upright transparent chamber 14, which is closedat its open end by removable cap 16. Cap 16 and chamber 14 are soconfigured at their junctures so that they define together a seal 18.The center of cap 16 is configured so as to define a receptacle 20.Receptacle 20 projects inwardly so that it opens into reaction chamber12 whencap 16 is in sealed relationship with chamber 14.

A sample support structure indicated generally at 22 comprises a shaft24 that is received in receptacle 20 at one end and is provided withtransversely extending slot 28 at its remote end. Porous member 26 isreceived in slot 28.

With particular reference to FIGS. 4 and 5, there is illustrated afilter holder 30 that is mounted to the closed exterior end of chamber14. Filter holder 30 receives and positions optical filter 32 so thatthe contents of reaction chamber 12 may be viewed through optical filter32. I

With particular reference to FIG. 6, there is illustrated a cap 34 thatreplaces cap 16 in sealed relationship with chamber 14. A hollow shaft36 is mounted in port 40 that extends axially through the center of cap34 from the exterior to the interior of reaction chamber 12. Theexterior; end of hollow shaft 36 is sealed with rubber seal 38, andporous member 26 is received in a slot in the innermost end of shaft 36.Hypodermic needle 42 is shown inserted through rubber seal 38 so as todischarge sample fluid into the interior of hollow shaft 36.

In use a body of liquid indicating reagent 46 is confined within thereaction chamber 12 so that liquid indicating reagent 46 rests on thecircular lower horizontal wall of chamber 14. The surface 48 of liquidindicating reagent 46 is exposed to the vapor phase that is confinedwithin reaction chamber 12. A sample of the body fluid that is beingtested for its volatile content is placed on porous member 26. Porousmember 26 im- -mediately expands to approximately 10 times its dryvolume so that the surface of the sample of body fluid is greatlyattenuated, thereby promoting the rapid vaporization of any volatilesubstance that may be in the liquid sample. The liquid indicatingreagent 46 is preferably swirled at predetermined time intervals so asto bring fresh reagent from the body of the liquid indicating reagent 46to the surface 48 where it will react with the volatiles in the vaporphase space within reaction iamber 12. The rounded lower corners 44 onreaction iamber 12 aid in swirling the liquid indicating reagent 6. Thelength of shafts '24 and 36 are adjusted so that hen porous member 26 isreceived in the transverse 5 ots in these respective shafts, it will besupported ithin reaction chamber 12 out of contact with the alls of thereaction chamber and liquid indicating rea- :nt 46. Only the vapor phaseis in common contact ith both the porous member 26 and the surface 48 ofuid indicating reagent 46. The spacing between the .rface 48 of liquidindicating reagent 46 and porous ember 26 is sufficient to preventliquid indicating rea- :nt 46 from coming into contact with porousmember 5 when liquid indicating reagent 46 is agitated by genswirling.In utilizing the analytic package 10 illustrated in FIG. the sample ofbody fluid which is to be tested for volile content is placed on poroussupport 26 while the lp 16 is separated from chamber 14. Cap 16 is thenimediately placed over chamber 14 so as to effect al 18. When utilizingthe analytic package 10 illustrated lrticularly in FIG. 6, the sample ofbody fluid is incted into the sealed reaction chamber 12 by insertingpodermic needle 42 through rubber seal 38 and forcg the sample of bodyfluid out of the hypodermic neee 42, down hollow shaft 36, onto porousmember 26. tilizing the embodiment particularly illustrated in G. 6, anyvolatile material that flashes into the vapor iase immediately on theexpansion of the surface area 'the liquid sample is trapped in reactionchamber 12. An optical filter, illustrated particularly in FIGS. 4 Id 5,is employed when color changes in the indicatg reagent are either notvisible to the unaided eye or e much easier to detect when observedthrough an itical filter having optical characteristics that areprelected for this purpose. Preferably, the surface 48 ofliquidindicating reagent i should be as large as possible in relation to thevolne of the vapor space in reaction chamber 12. The mple of body fluidis generally quite small, being usuy not larger than one-half of amilliliter. If the vapor lase space in reaction chamber 12 is large, thevolae component that is volatilized from the liquid samon porous member26 is diluted to such an extent at the reaction with the liquidindicating reagent 46 unreliable. In a preferred embodiment the exteriordiameter of amber 14 is 27 millimeters, and the total exterior ight withcap 16 in place is 32 millimeters. The disice from the inwardly disposedside of cap 16 to the note end of shaft 24 is 18 millimeters; thediameter shaft 24 is 2 millimeters; slot 28 is l millimeter wide d 5millimeters deep; porous member 26 is 10 milli- :ters long, 6millimeters wide, and l millimeter thick; d the transparent walls ofchamber 14 are about 1 llimeter thick. ?referably, analytic package 10is proportioned so it for every square centimeter of surface 48, thereis m about 2 to 3 cubic centimeters of vapor phase ace within reactionchamber 12; however, the ratio liquid reagent surface area to vaporphase volume ty range up to as much as 1 square centimeter to 5 morecubic centimeters. In general, the proportion surface area to vaporspace volume in reaction chamber 12 should be no greater thanapproximately 1 square centimeter to 5 or 6 cubic centimeters.

The analytical package and method described are particularly suited forrapidly detecting qualitatively and semiquantitatively ethanol,methanol, and isopropanol in blood, urine, and cerebral spinal fluid.

The surface area of the sample of fluid may be augmented or expanded topromote vaporization of its volatile components through the use ofvarious porous materials. Suitable sample surface area expandersinclude, for example, clays, cotton, elemental carbon, openporecellulosic foam, and the like. Preferably, the openpore cellulosic foammaterial is used because it is inert with respect to the body fluidsamples, and it expands so that the surface area of the sample is vastlyexpanded with great rapidity. In general, it is necessary to use largersamples of body fluid when porous support members other than cellulosicfoam are employed. The increased volume of the sample compensates forthe decrease in sample surface area expansion. In general, the volatilecomponent, the presence of which is to be detected, is present in thesample. It is possible, however, to provide a reagent on the porousmember such that a reaction between the component being tested for inthe sample and the reagent on the porous member will generate a volatilesubstance that may be detected by the indicating reagent.

In general, the most convenient form for the analytic package is that ofan upright cylinder. If desired, however, the analytic package may beconfigured to meet the requirements of a particular usage. For example,if the analytic package is to be used in a spectrophotometer, it may benecessary to configure the analytic package as a square or rectangularcolumn.

The liquid indicating reagent is preferably chosen so that upon reactionwith the volatile material being tested for it will give a visuallydetectable indication. Suitable liquid indicating reagents include; forexample, admixtures of sulfuric acid and potassium permanganate andadmixtures of sulfuric acid and potassium dichromate.

In one procedure applied to detect the presence and concentration ofethanol in blood, two separate reagent solutions are prepared. Asulfuric acid solution is prepared by admixing 1 part of concentratedsulfuric acid with 2 parts of water. A dilute potassium permanganatesolution is prepared by admixing potassium permanganate with water toproduce a 0.02 Normal potassium permanganate solution. One milliliter ofthe potassium permanganate solution is diluted to 20 milliliters withthe sulfuric acid solution. A microdiffusion chamber having thedimensionsdescribed above is selected for use as the reaction chamber,and 2 milliliters of the freshly prepared potassium permanganatesulfuric acid solution is added to the chamber. A 0.1 milliliter sampleof .blood is applied to the cellulosic foam support member, and the capis screwed into place on the chamber so that a vapor-tight seal betweenthe cap and the chamber is achieved. The chamber is swirled gently,without allowing contact between the indicating reagent and the supportmember. It is allowed to stand for 1 minute and reswirled at 1 minuteintervals for a period of 4 minutes. The disappearance of thepermanganate color indicates the presence of a volatile reducingsubstance. Repetition of this procedure using cotton as the porousmember produces the same indication when the blood sample size isincreased to approximately 1 milliliter.

The indication of ethanol in blood using this procedure is such that thetime required for the disappearance of the permanganate color isdirectlyproportional to the concentration of ethanol. Thus, if the colordisappears in exactly 1 minute, the ethanol concentration isapproximately 0.5 grams perlOO milliliters of blood. If the permanganatecolor disappears in 5 minutes, the concentration of ethanol in the bloodis approximately 0.10 grams per 100 milliliters ofblood.

The apparatus and procedure described herein are generally applicable toanalytical problems where a volatile component in an admixture ofliquids must be detected qualitatively and semiquantitatively in onesimple, quick step. In addition to the analysis of botanical andbiological fluids, the procedure herein is applicable to variousmanufacturing procedures. For example, in solution polymerization wherea volatile monomer is employed, it is possible to detect the progress ofthe reaction and the end point of the polymerization reaction byobserving the decrease in the concentration of the volatile monomer overa period of time. Successive samples are withdrawn from the reactionvessel and are subjected to analysis, as described herein, with thedecrease in concentration of the monomer being detectable by increasingreaction times in the reaction chamber.

The reagents and sample are chosen so that the indication is obtained atnormal room temperatures. If an additional heating step is required tovolatilize the volatile component, the chances for error aresubstantially multiplied and the objective of simplicity is defeated.Volatilization of the liquid sample is achieved by greatly expanding itssurface area rather than by heating it. In general, the surface area ofthe liquid sample is expanded to an area at least times and preferablytimes greater than that of the same sample in spherical form so as topromote rapid volatilization at room temperature. Also, theproportioning of the surface area of the liquid indicating reagent tothe volume of the vapor space is chosen so that a rapid, reliableindication is given by the reagent. In general, when the analyticpackage in its assembled form is substantially a right cylinder, thereaction chambers height should be least than approximately three timesits diameter.

What is claimed is:

1. An analytical method comprising:

expanding the surface area of a liquid sample;

confining the resultant expanded sample within a closed chamber, saidclosed chamber also containing a vapor phase space and a liquidindicating reagent, said expanded sample being positioned in closeproximity to and out of contact with said liquid indicating reagent;

allowing any volatile substance present with said expanded sample tovaporize to about room temperature into said vapor phase space and toreact with said liquid indicating reagent; and

observing said liquid indicating reagent for a predetermined period oftime to detect any indication of a reaction with said volatilesubstance.

2. An analytical method of claim 1 wherein the surface area of theliquid sample is expanded to an area at least 20 times greater than thatof the same sample in spherical form.

- 3 Ananalytical method of claim 1 including observing the indicatingreagent to detect any change in color in said indicating reagent.

4. An analytical method of claim 1 including placing the liquid sampleon a cellulosic foam.

5. An analytical method comprising:

placing a sample of body fluid on a porous support member to expand thesurface area of said sample;

confining the vapor phase surrounding the resultant expanded sample in aclosed, substantially transparent, reaction chamber; providing liquidindicating reagent'in said reaction chamber, said indicating reagent andsaid expanded sample both being in contact with said vapor phase and outof contact with one another; allowing any volatile substance present insaid expanded sample to vaporize into said vapor phase and to react withsaid indicating reagent; and observing said indicating reagent for apredetermined period of time and agitating said indicating reagent atpredetermined intervals during said period of time to detect anyindication of the reaction of said indicating reagent with said volatilesubstance.

6. An analytic method of claim 5 including placing a sample of blood ona porous support member, providing an aqueous admixture of potassiumpermanganate and sulfuric acid in the reaction chamber, allowing anyalcohol present in the expanded sample to vaporize,

and observing said aqueous potassium permanganate sulfuric acid solutionfor a period of time to detect the disappearance of the permanganatecolor.

7. An analytical package comprising:

closable reaction chamber means enclosing within a common vapor phasespace, a sample support means, and an indicating reagent, said samplesupport means and a generally liquid indicating reagent being spaced inclose proximity to and out of contact with one another; and

said sample support means including a porous member for supporting aliquid sample and extending the surface area of said liquid sample to anarea at least 10 times greater than that of the samesample in sphericalform to promote the vaporization of a volatile substance from saidliquid sample on said porous member, said indicating reagent beingadapted to give a detectable indication responsive to the presence ofsaid volatile substance in said vapor phase, the ratio of the surfacearea of said generally liquid indicating reagent to the volume of saidcommon vapor phase space being in the proportion of 1 square centimeterto no more than approximately 6 cubic centimeters.

8. An analytical package of claim 7 wherein at least one wall of theclosable reaction chamber means is substantially transparent.

9. An analytical package of claim 7 including means for permitting theliquid sample to be placed on the porous member while the closablereaction chamber means is in the closed configuration.

10. An analytical package of claim 7 wherein the porous member is inertto the liquid sample.

1 1. An analytical package of claim 7 wherein the porous member is acellulosic foam.

12. An analytical package of claim 7 including an optical filterpositioned so as to permit viewing of the indicating reagent throughsaid optical filter.

13. An analytical package comprising:

cover member within said reaction chamber, said means for mountingpositioning said porous memher at a location spaced from said closed enda distance sufficient to permit a body of liquid to be positioned onsaid closed end within said reaction chamber out of contact with saidporous member, said reaction chamber being proportioned so that theratio of the surface area of said body of liquid to the volume of thevapor space within said chamber is in the proportion of 1 squarecentimeter to as much as 5 cubic centimeters.

1. AN ANALYTICAL METHOD COMPRISING: EXPANDING THE SURFACE AREA OF ALIQUID SAMPLE; CONFINING THE RESULTANT EXPANDED SAMPLE WITHIN A CLOSEDCHAMBER, SAID CLOSED CHAMBER ALSO CONTAINING A VAPOR PHASE SPACE AND ALIQUID INDICATING REAGENT, SAID EXPANDED SAMPLE BEING POSITIONED INCLOSE PROXIMITY TO AND OUT OF CONTACT WITH SAID LIQUID INDICATINGREAGENT; ALLOWING ANY VOLATILE SUBSTANCE PRESENT WITH SAID EXPANDEDSAMPLE TO VAPORIZE TO ABOUT ROOM TEMPERATURE INTO SAID VAPOR PHASE SPACEAND TO REACT WITH SAID LIQUID INDICATING REAGENT; AND OBSERVING SAIDLIQUID INDICATING REAGENT FOR A PREDETERMINED PERIOD OF TIME TO DIRECTANY INDICATION OF A REACTION WITH SAID VOLATILE SUBSTANCE.
 2. Ananalytical method of claim 1 wherein the surface area of the liquidsample is expanded to an area at least 20 times greater than that of thesame sample in spherical form.
 3. An analytical method of claim 1including observing the indicating reagent to detect any change in colorin said indicating reagent.
 4. An analytical melthod of claim 1including placing the liquid sample on a cellulosic foam.
 5. Ananalytical method comprising: placing a sample of body fluid on a poroussupport member to expand the surface area of said sample; confining thevapor phase surrounding the resultant expanded sample in a closed,substantially transparent, reaction chamber; providing liquid indicatingreagent in said reaction chamber, said indicating reagent and saidexpanded sample both being in contact with said vapor phase and out ofcontact with one another; allowing any volatile substance present insaid expanded sample to vaporize into said vapor phase and to react withsaid indicating reagent; and observing said indicating reagent for apredetermined period of time and agitating said indicating reagent atpredetermined intervals during said period of time to detect anyindication of the reaction of said indicating reagent with said volatilesubstance.
 6. An analytic method of claim 5 including placing a sampleof blood on a porous support member, providing an aqueous admixture ofpotassium permanganate and sulfuric acid in the reaction chamber,allowing any alcohol present in the expanded sample to vaporize, andobserving said aqueous potassium permanganate sulfuric acid solution fora period of time to detect the disappearance of the permanganate color.7. An analytical package comprising: closable reaction chamber meansenclosing within a common vapor phase space, a sample support means, andan indicating reagent, said sample support means and a generally liquidindicating reagent being spaced in close proximity to and out of contactwith one another; and said sample support means including a porousmember for supporting a liquid sample and extending the surface area ofsaid liquid sample to an area at least 10 times greater than that of thesame sample in spherical form to promote the vaporization of a volatilesubstance from said liquId sample on said porous member, said indicatingreagent being adapted to give a detectable indication responsive to thepresence of said volatile substance in said vapor phase, the ratio ofthe surface area of said generally liquid indicating reagent to thevolume of said common vapor phase space being in the proportion of 1square centimeter to no more than approximately 6 cubic centimeters. 8.An analytical package of claim 7 wherein at least one wall of theclosable reaction chamber means is substantially transparent.
 9. Ananalytical package of claim 7 including means for permitting the liquidsample to be placed on the porous member while the closable reactionchamber means is in the closed configuration.
 10. An analytical packageof claim 7 wherein the porous member is inert to the liquid sample. 11.An analytical package of claim 7 wherein the porous member is acellulosic foam.
 12. An analytical package of claim 7 including anoptical filter positioned so as to permit viewing of the indicatingreagent through said optical filter.
 13. An analytical packagecomprising: a substantially transparent reaction chamber being closed atone end and open at the other end, the height of said reaction generallybeing less than approximately three times the diameter of said reactionchamber; a cover member for closing said open end; a porous member forsupporting a liquid sample and extending the surface area of said liquidsample to an area at least 10 times greater than the surface area of thesame liquid sample in spherical form; and means for mounting said porousmember on said cover member within said reaction chamber, said means formounting positioning said porous member at a location spaced from saidclosed end a distance sufficient to permit a body of liquid to bepositioned on said closed end within said reaction chamber out ofcontact with said porous member, said reaction chamber beingproportioned so that the ratio of the surface area of said body ofliquid to the volume of the vapor space within said chamber is in theproportion of 1 square centimeter to as much as 5 cubic centimeters.